We recently published in Nature Medicine that low nitric oxide (NO) bioavailability contributes to cerebral malaria (CM), the neurological disorder that kills over 2 million people annually;thus, improving NO bioavailability represents a potential adjunctive therapy for this deadly disease. Adjunctive therapy is important because about 30% of CM patients succumb despite appropriate anti-parasite chemotherapy. Low NO bioavailability in experimental cerebral malaria contributes to inflammation and vascular leak during experimental cerebral malaria (ECM). In other diseases, low NO bioavailability contributes to platelet activation and aggregation, endothelial dysfunction, vascular leak, and immune activation and regulation;these events also contribute to the complex malaria syndrome. NO therapy may therefore be effective against this complex disease because of its pleiotropic inhibitory effects of the above processes. Indeed, inhaled NO and i.v. injection of a NO donor significantly (P<0.05) protected against ECM mortality, providing the rationale for being a potential adjunctive therapy for CM. Unfortunately, neither of the above NO therapies are suitable for implementation in resource poor clinics in malaria endemic areas. There are, however, alternate approaches to restore NO bioavailability during ECM that will work in these clinics. Because the low NO bioavailability during CM is caused in part by severe hypoargininemia, which leads to decreased endothelial nitric oxide synthase-derived NO production, one approach is to restore arginine levels by parenteral administration of the amino acid. This is the basis for Anstey and colleagues ongoing trials that have shown marked restoration of NO-dependent endothelial cell function during human falciparum malaria. We posit that citrulline will be a more effective approach to restore arginine levels and hence NO bioavailability because (i) citrulline inhibits arginases released from RBCs during malaria, (ii) citrulline is not metabolized by the liver, (iii) citrulline is less toxic than arginine, and (iv) citrulline ameliorates hypoargininemia better than arginine itself. Citrulline is converted by the kidneys into arginine. Indeed, our preliminary indicate that citrulline completely protects all recipients against ECM mortality whereas the same dose of arginine caused toxicity and early death. To provide the initial proof of concept required to move toward an IND, we propose in aim 1 to measure the pharmacokinetics of arginine and citrulline therapy. In aim 2, we compare the efficacy of equimolar arginine and citrulline in protecting against mortality during ECM. Because mortality by citrulline and arginine therapy can only be improved by decreasing ECM pathogenesis and restoring NO production, we will confirm the differences in mortality with pathogenesis and NO bioavailability studies. Aim 3 determines whether the adjunctive therapy affects anti-parasite chemotherapy. Based on the complete protection against ECM by citrulline and the studies in the cardiovascular and sickle cell disease fields, we anticipate that citrulline represents a novel IND for cerebral malaria, a life threatening neurological disorder. As such, the proposed therapy meets the guidelines for the R21 Exploratory/developmental Projects in Translation Research, which is designed to test new therapeutics with mechanisms not as yet supported by substantial data and precedent. Public Health Relevance: We have reported the low nitric oxide bioavailability contributes to the pathogenesis of experimental cerebral malaria. This research finding is translated into a potential adjunctive therapy for cerebral malaria that rescues patients from their neurological disease. The proposed research generates proof-of-principle data needed to move the proposed adjunctive therapy toward an Investigational New Drug application. Disclaimer: Please note that the following critiques were prepared by the reviewers prior to the Study Section meeting and are provided in an essentially unedited form. While there is opportunity for the reviewers to update or revise their written evaluation, based upon the group's discussion, there is no guarantee that individual critiques have been updated subsequent to the discussion at the meeting. Therefore, the critiques may not fully reflect the final opinions of the individual reviewers at the close of group discussion or the final majority opinion of the group. Thus the Resume and Summary of Discussion is the final word on what the reviewers actually considered critical at the meeting.